L.lactis KLDS4.0325产细菌素发酵培养基的响应面优化

Optimization of fermentation medium for bacteriocin production of L.lactis KLDS4.0325 by response surface methodology

以分离自新疆牧民家庭自制酸马奶中的乳酸乳球菌KLDS4.0325为研究对象,以大肠杆菌为指示菌,以抑菌圈直径为考察指标,通过单因素实验(碳源、氮源及复合氮源)、Plackett-Burman、最陡爬坡实验和Box-Behnken实验优化其产细菌素培养基。PB实验结果表明,对细菌素产量影响显著的因素有蔗糖、蛋白胨、抗坏血酸钠;经Box-Behnken实验优化后培养基配方为:128.99 g/L蔗糖,17.59 g/L蛋白胨,10 g/L酪蛋白胨,10 g/L磷酸氢二钾,60 g/L碳酸钙,1.5 g/L硫酸镁,1.49 g/L抗坏血酸钠,1.5 g/Lβ-甘油磷酸钠,乙酸钠、氯化钠、硫酸锰各为1.0 g/L;该条件下抑菌圈直径为24.76 mm,效价可达5000 U/m L,比优化前提高了3.31倍。并经验证实验证明该配方下的实验结果与预测值吻合,因此该回归模型是切实可行的。

The fermentation medium composition of L.Lactis KLDS4.0325 isolated from self-made koumiss Xinjiang were optimized for bacteriocin production by single factor experiment （ carbon source, nitrogen source, different mixed nitrogen sources）, Plackett- Burman, The steepest grade test and Box- Behnken design double- plate method.The inhibitory zone diameter（mm）was used as evaluation index and quantified against Escherichia coil ATCC 25922.PB Experiment results showed that sucrose,peptone and sodium ascorbate had significant influence on bacteriocin production.And finally the optimal combination of the medium constituents for bacteriocin production was determined as： 128.99 g/L sucrose, 17.59 g/L peptone, 10 g/L casein peptone, 10 g/L K2HPO4,60 g/L CaCQ, 1.5 g/L MgSO4 -7 H2 O, 1.49 g/L sodium ascorbate, 1.5 g/L/3- glycerophosphate, CH3 COONa. 3 H2 O, NaCI, MnSO4 respectively L0 g/L.Under the optimal culture condition,the inhibitory zone diameter was 2426 mm and the bacteriocin inhibitory activity was increased by 3.31 times and reached up to 5000 U/mL.Meanwhile,the consistent results between the prediction and experiments indicated the established model in this study is feasible.